Project Summary The liver plays a pivotal role in the maintenance of lipid homeostasis. Accumulation of lipids in the liver with no alcoholic consumption leads to nonalcoholic fatty liver diseases (NAFLD). NAFLD spectrum ranges from a simple nonalcoholic fatty liver (steatosis) to steatohepatitis (NASH) and cirrhosis. Insulin resistant metabolic disorder and subsequent inflammation are the major pathogenesis for development of the diseases, but the details of the mechanisms are not fully understood. Most studies have focused on the role of free fatty acid-mediated lipotoxicity. It has however been shown that there is widespread dysregulation of lipid metabolism in NAFLD and, specifically, there are major perturbations in cholesterol metabolism. The potential mechanisms by which such perturbations may lead to liver diseases remain unknown. This gap in the field is a major barrier towards understanding the role of cholesterol metabolites in the pathogenesis of NASH and leveraging this information to develop novel therapies for NAFLD. We recently identified novel regulatory cholesterol metabolites, 25HC3S and 25HCDS, in normal human plasma and liver tissues. Administration of the metabolites or increase in expression of hydroxysterol sulfotransferase (SULT2B1b, the key enzyme for the synthesis of these metabolites) decreases lipid accumulation in serum and liver tissues. This proposal focuses on the role of the metabolites in the development of NAFLD. Hypothesis: Based on our strong preliminary data, we propose that decreases in SULT2B1b activity with consequent decreases in 25HC3S/25HCDS promote hepatic steatosis by (1) fail to inhibit the pro-lipogenic transcriptional factor sterol response element binding proteins (SREBPs) and (2) decrease expression and activity of the peroxisome proliferator- activated receptor-? (PPAR?) via epigenomic regulation, activating histone deacetylase (SirT1). We will test the hypothesis by the following specific aims: 1) To elucidate the biochemical mechanism by which the cholesterol metabolites regulate lipid metabolism and inflammatory responses. 2) To explore the role of SULT2B1b in development of NALFD in vitro in free fatty acid-induced and in vivo high fat diet-induced NALFD models. 3) To test the potential of the cholesterol metabolites to prevent/reverse hepatic lipid accumulation in NAFLD animal models. The overall goal of this proposal is to understand the molecular mechanisms of oxysterol sulfation involved in the coordinate regulation of hepatocyte lipid metabolism and inflammatory responses, and to explore its potential clinical utility as a treatment for NAFLD.